RESUMO
Bioactivity-guided chromatographic methods are of great significance for the isolation of the active compounds in complex samples. In this study, four anti-fungal compounds were located by activity screening and successfully isolated from a microbial fermentation sample by preparative high-performance liquid chromatography. Separation performance of columns including C18, positively charged C18, negatively charged C18 and C8 were firstly investigated. And it showed a better capacity of mixed-mode stationary phases for retention and separation. Therefore, the positively charged C18 column was used to separate the sample into several fractions, among which the active one was identified by the antifungal test. And then the active fraction was enriched and separated again by successively using the negatively charged C18 and C8 columns to obtain four compounds, which were identified as polyoxins A, K, F and H. With activity verification, four polyoxins were found to have good inhibitory effects against the three fungal plant diseases including rice sheath blight, tomato grey mould disease, and apple spot leaf disease.
Assuntos
Antifúngicos/isolamento & purificação , Antifúngicos/química , Antifúngicos/metabolismo , Cromatografia Líquida de Alta Pressão , Fermentação , Estrutura Molecular , Nucleosídeos de Pirimidina/química , Nucleosídeos de Pirimidina/isolamento & purificação , Nucleosídeos de Pirimidina/metabolismoRESUMO
The chromatographic behavior of a kind of nucleoside peptides, polyoxins, was investigated in this study. Molecular simulation technique was used to elucidate the temperature-dependent peak sharpening of polyoxins. There was a relatively small energy barrier between the global minimum conformer and the local minimum conformer of polyoxin A and the high temperature helped to quickly cross the energy barrier and accelerate the conformational transformation for getting the global minimum, so that stationary phase could not identify these two conformations and presented a sharp peak. Two kinds of mixed-mode columns, strong cation exchange or strong anion exchange ligands bonded with C18 (C18SCX and C18SAX) were used to improve separation selectivity of four polyoxins (A, K, F, H). The electrostatic attraction was necessary to increase the retention to ensure that the alkyl chain can give better play to its hydrophobic effect. Therefore, four polyoxins were well separated on C18SCX at pH 2 and they were also well separated on C18SAX at pH 7. In the small-scale purification of polyoxins, the sample loading of the C18SCX was five times than that of the C18SAX and the purity of the collected four polyoxins was all over 90%.
Assuntos
Cromatografia Líquida de Alta Pressão , Concentração de Íons de Hidrogênio , Conformação Molecular , Simulação de Dinâmica Molecular , Nucleosídeos de Pirimidina/química , Nucleosídeos de Pirimidina/isolamento & purificação , Sais/químicaRESUMO
An organic-silica hybrid monolith was prepared by a single-step ring-opening polymerization of octaglycidyldimethylsilyl polyhedral oligomeric silsesquioxane (POSS-epoxy), polyethylenimine (PEI), and ß-cyclodextrin (ß-CD) in a ternary porogenic solvent consisting of polyethylene glycol, 1,4-butanediol, and 1-propanol. The framework of POSS-PEI hybrid monolith could offer well-defined 3D skeleton, while ß-CD with the ability of forming a host-guest inclusion complexes with a variety of compounds could show an ability of specific selection. The obtained hybrid monoliths were successfully applied for separation of phenols, benzoic acids, and nucleobases. Especially due to the introduction of ß-CD, positional isomers including hydroquinone and resorcinol, o-nitrophenol and p-nitrophenol, as well as p-chlorophenol and o-chlorophenol were baseline separated and the column efficiency reached 82 300 plates/m for hydroquinone.
Assuntos
Eletrocromatografia Capilar/instrumentação , Compostos de Epóxi/química , Compostos de Organossilício/química , Benzoatos/análise , Benzoatos/química , Benzoatos/isolamento & purificação , Eletrocromatografia Capilar/métodos , Fenóis/análise , Fenóis/química , Fenóis/isolamento & purificação , Polimerização , Nucleosídeos de Pirimidina/análise , Nucleosídeos de Pirimidina/química , Nucleosídeos de Pirimidina/isolamento & purificação , beta-Ciclodextrinas/químicaRESUMO
Eleven new pyrimidine nucleosides (1-11) and 12 known analogues (12-23) were isolated from the marine-derived Streptomyces sp. SSA28. All of the new structures were elucidated by extensive NMR spectroscopic analysis and HRESIMS data. The absolute configurations of compound 1 were determined by X-ray diffraction. The configurations of 2-16 were investigated by ECD calculations. Compounds 11-16 showed cytotoxicity against HCT-116 human colon cancer cell lines with IC50 values from 0.39 ± 0.03 to 6.63 ± 0.47 µM.
Assuntos
Nucleosídeos de Pirimidina/isolamento & purificação , Streptomyces/química , Cristalografia por Raios X , Ensaios de Seleção de Medicamentos Antitumorais , Células HCT116 , Humanos , Estrutura Molecular , Nucleosídeos de Pirimidina/química , Análise Espectral/métodosRESUMO
Two approaches for C4 modifications of silyl-protected thymidine, 2'-deoxyuridine, and 3'-azido-2',3'-dideoxythymidine (AZT) are described. In both, nucleoside amide activation with 1H-benzotriazol-1-yloxy-tris(dimethylamino)phosphonium hexafluorophosphate (BOP) and DBU yields O4 -(benzotriazol-1-yl) derivatives. These in situ-formed intermediates are reacted with various nucleophiles, resulting in C4 modifications. In the two-step, one-pot approach, the O4 -(benzotriazol-1-yl) nucleoside intermediates are initially produced by reactions of the nucleosides with BOP and DBU in THF. This step is fast and typically complete within 30 min. Subsequently, the O4 -(benzotriazol-1-yl) derivatives are reacted with nucleophiles, such as aliphatic and aromatic amines, thiols, and alcohols, under appropriate conditions. Workup, isolation, and purification lead to the desired C4-modified pyrimidine nucleosides in good to excellent yields. In the one-step approach, the nucleosides are reacted with BOP and DBU, in the presence of the nucleophile (only aliphatic and aromatic amines, and thiols have been tested). Where comparisons are possible, the one-step approach is generally superior. © 2019 by John Wiley & Sons, Inc.
Assuntos
Amidas/química , Fluoretos/química , Fosfatos/química , Nucleosídeos de Pirimidina/química , Espectroscopia de Ressonância Magnética Nuclear de Carbono-13 , Cromatografia em Gel , Cromatografia em Camada Fina , Espectroscopia de Prótons por Ressonância Magnética , Nucleosídeos de Pirimidina/isolamento & purificação , Espectrometria de Massas por Ionização por Electrospray , Espectrofotometria UltravioletaRESUMO
In the course of our screening for selective growth inhibitors of human umbilical vein endothelial cells (HUVECs), we isolated sangivamycin from the culture filtrate of Streptomyces. It inhibited the growth of HUVECs at approximately 30 times lower concentration than that needed to inhibit the growth of WI-38 human fibroblasts. Structurally-related nucleosides, such as toyocamycin, tubercidin, and formycins A and B, did not show the differential inhibition. Although sangivamycin is known to inhibit protein kinase C, other protein kinase C inhibitors did not inhibit the growth of HUVECs selectively. Sangivamycin effectively inhibited S-phase induction in HUVECs, like TNP-470 and LLnL, known selective inhibitors. However, unlike them sangivamycin did not induce p21 expression. On the other hand, sangivamycin was found to inhibit DNA synthesis selectively in HUVECs. Thus, sangivamycin was shown to be a new selective growth inhibitor of HUVECs acting on DNA synthesis.
Assuntos
Antibióticos Antineoplásicos/farmacologia , Endotélio Vascular/efeitos dos fármacos , Nucleosídeos de Pirimidina/farmacologia , Western Blotting , Divisão Celular/efeitos dos fármacos , Células Cultivadas , Colágeno/química , Relação Dose-Resposta a Droga , Combinação de Medicamentos , Fibroblastos/efeitos dos fármacos , Humanos , Laminina/química , Inibidores da Síntese de Ácido Nucleico/farmacologia , Proteoglicanas/química , Nucleosídeos de Pirimidina/isolamento & purificação , Streptomyces/metabolismo , Timidina/química , Veias Umbilicais/citologiaRESUMO
A novel nucleoside antibiotic, SF2457, was isolated from the fermentation broth of Nocardia brasiliensis SF2457. The structure of SF2457 was determined by degradation studies using alkaline hydrolysis and methanolysis. SF2457 is closely related to the amicetin group antibiotics. The antibiotic exhibited inhibitory activity against Gram-positive and Gram-negative bacteria.
Assuntos
Antibacterianos/isolamento & purificação , Nocardia/metabolismo , Antibacterianos/biossíntese , Antibacterianos/química , Fermentação , Nucleosídeos de Pirimidina/química , Nucleosídeos de Pirimidina/isolamento & purificação , Nucleosídeos de Pirimidina/farmacologiaRESUMO
A bacterial isolate from soil, designated as TB-2125 had a unique pattern of multiple resistance to aminoglycoside antibiotics (AG) and produced new nucleoside antibiotics. Taxonomic properties of this strain fell into those of Bacillus circulans, providing unique characteristics such as strict susceptibility to acidic pH, motility of colony as well as multiple AG-resistance. Two new antibiotics which were named bagougeramines A and B had a broad antimicrobial activity and a specific activity against the two spotted spider mite.
Assuntos
Antibacterianos/isolamento & purificação , Bacillus/classificação , Animais , Antibacterianos/farmacologia , Bacillus/metabolismo , Fermentação , Camundongos , Ácaros/efeitos dos fármacos , Nucleosídeos de Pirimidina/isolamento & purificação , Nucleosídeos de Pirimidina/farmacologiaRESUMO
A novel class of antibiotics was isolated from cultures of Streptomyces coeruleorubidus strain AB 1183F-64. The antimicrobial activity of the pacidamycins is selective against Pseudomonas aeruginosa. The various congeners are nucleoside peptides which differ in the terminal amino acid residues. The structures were determined using MS-MS and 2D NMR techniques.
Assuntos
Antibacterianos/análise , Oligopeptídeos/análise , Peptídeos , Pseudomonas aeruginosa/efeitos dos fármacos , Microbiologia do Solo , Streptomyces/metabolismo , Uridina/análogos & derivados , Antibacterianos/isolamento & purificação , Concentração de Íons de Hidrogênio , Peptídeos e Proteínas de Sinalização Intercelular , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Estrutura Molecular , Oligopeptídeos/isolamento & purificação , Nucleosídeos de Pirimidina/análise , Nucleosídeos de Pirimidina/isolamento & purificação , SolubilidadeRESUMO
Development of a liquid chromatographic method which can separate each of a series of hexopyranosylated cytosine nucleosides from their degradation products formed at acid, neutral and basic pH is described. Both silica-based reverse-phase and polymer columns were examined. Influence of the mobile phase pH, ion-pairing agent, concentration of the buffer and type and concentration of organic modifier were systematically investigated. The concentration of the ion-pairing agent and the buffer were found to have a major effect on selectivity. Samples were finally analyzed on a poly(styrene-divinylbenzene), PLRP-S 100 A (8 microns) 250 x 4.6 mm I.D. column at 60 degrees C and with a mobile phase consisting of acetonitrile-sodium octanesulphonate (pH 2.5; 0.02 M)-potassium phosphate buffer (pH 2.5; 0.2 M)-water (X:25:50:25-X, v/v, where X is variable).
Assuntos
Antineoplásicos/isolamento & purificação , Antivirais/isolamento & purificação , Citosina/isolamento & purificação , Nucleosídeos de Pirimidina/isolamento & purificação , Antineoplásicos/química , Antivirais/química , Cromatografia Líquida , Citosina/análogos & derivados , Citosina/química , Concentração de Íons de Hidrogênio , Indicadores e Reagentes , Nucleosídeos de Pirimidina/química , Análise de RegressãoRESUMO
Syntheses of specified 2'-modified nucleosides were achieved: a) via oximation of the 5',3'-blocked 2'-oxocytidine, followed by reduction, or b) by intramolecular nucleophilic addition of 3'-(2-methoxyethoxy)carbamate to the 2'-position with opening of O(2),2'-anhydrouridine. For the first time, 3'-phosphoroamidites of these 2'-modified nucleosides were successfully incorporated into oligonucleotides by solid-phase synthesis. Incorporation of 2'-modified nucleotides into oligodeoxyribonucleotides had a negative effect on the duplex T(m) values with the DNA or RNA complements. Nevertheless, modified nucleotides have shown good target recognition; the (S)-isomer binds preferably to RNA and the (R)-isomer to DNA. Both modified nucleosides significantly increased nuclease resistance of the oligodeoxyribonucleotides.
Assuntos
Oligonucleotídeos/síntese química , Nucleosídeos de Pirimidina/síntese química , Conformação de Ácido Nucleico , Oligonucleotídeos/isolamento & purificação , Persea , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Nucleosídeos de Pirimidina/isolamento & purificação , Venenos de Serpentes/química , Venenos de Serpentes/isolamento & purificação , EstereoisomerismoRESUMO
Improved nitrogen utilization in cattle is important in order to secure a sustainable cattle production. As purines and pyrimidines (PP) constitute an appreciable part of rumen nitrogen, an improved understanding of the absorption and intermediary metabolism of PP is essential. The present work describes the development and validation of a sensitive and specific method for simultaneous determination of 20 purines (adenine, guanine, guanosine, inosine, 2'-deoxyguanosine, 2'-deoxyinosine, xanthine, hypoxanthine), pyrimidines (cytosine, thymine, uracil, cytidine, uridine, thymidine, 2'-deoxyuridine), and their degradation products (uric acid, allantoin, ß-alanine, ß-ureidopropionic acid, ß-aminoisobutyric acid) in blood plasma of dairy cows. The high performance liquid chromatography-based technique coupled to electrospray ionization tandem mass spectrometry (LC-MS/MS) was combined with individual matrix-matched calibration standards and stable isotopically labelled reference compounds. The quantitative analysis was preceded by a novel pre-treatment procedure consisting of ethanol precipitation, filtration, evaporation and reconstitution. Parameters for separation and detection during the LC-MS/MS analysis were investigated. It was confirmed that using a log-calibration model rather than a linear calibration model resulted in lower CV% and a lack of fit test demonstrated a satisfying linear regression. The method covers concentration ranges for each metabolite according to that in actual samples, e.g. guanine: 0.10-5.0 µmol/L, and allantoin: 120-500 µmol/L. The CV% for the chosen quantification ranges were below 25%. The method has good repeatability (CV%≤25%) and intermediate precision (CV%≤25%) and excellent recoveries (91-107%). All metabolites demonstrated good long-term stability and good stability within-runs (CV%≤10%). Different degrees of absolute matrix effects were observed in plasma, urine and milk. The determination of relative matrix effects revealed that the method was suitable for almost all examined PP metabolites in plasma drawn from an artery and the portal hepatic, hepatic and gastrosplenic veins and, with a few exceptions, also for other species such as chicken, pig, mink, human and rat.
Assuntos
Nucleosídeos de Purina/isolamento & purificação , Nucleosídeos de Pirimidina/isolamento & purificação , Espectrometria de Massas em Tandem/normas , Animais , Calibragem , Bovinos , Cromatografia Líquida de Alta Pressão , Humanos , Leite/química , Vison , Nucleosídeos de Purina/sangue , Nucleosídeos de Purina/urina , Nucleosídeos de Pirimidina/sangue , Nucleosídeos de Pirimidina/urina , Ratos , Padrões de Referência , Espectrometria de Massas por Ionização por Electrospray , SuínosAssuntos
Imidazóis , Nucleosídeos de Pirimidina/isolamento & purificação , Adenosina , Sítios de Ligação , Cromatografia Gasosa , Cromatografia em Camada Fina , Citosina , Desoxirribonucleosídeos/isolamento & purificação , Estudos de Avaliação como Assunto , Floxuridina , Guanosina , Isomerismo , Cinética , Métodos , Conformação Molecular , Peso Molecular , Conformação de Ácido Nucleico , Dióxido de Silício , Espectrofotometria Ultravioleta , Relação Estrutura-Atividade , Temperatura , Fatores de Tempo , UridinaAssuntos
Antibacterianos/biossíntese , Nucleosídeos de Pirimidina/análise , Meios de Cultura/análise , Fermentação , Guanidinas/biossíntese , Ácidos Hexurônicos/análise , Ácidos Hexurônicos/biossíntese , Ácidos Hexurônicos/isolamento & purificação , Conformação Molecular , Nucleosídeos , Nucleosídeos de Pirimidina/biossíntese , Nucleosídeos de Pirimidina/isolamento & purificação , Streptomyces/metabolismoAssuntos
Antinematódeos/isolamento & purificação , Compostos Organofosforados/isolamento & purificação , Nucleosídeos de Pirimidina/isolamento & purificação , Streptomyces/química , Animais , Antinematódeos/química , Antinematódeos/farmacologia , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Nematoides/efeitos dos fármacos , Compostos Organofosforados/química , Compostos Organofosforados/farmacologia , Nucleosídeos de Pirimidina/química , Nucleosídeos de Pirimidina/farmacologiaAssuntos
Antibióticos Antineoplásicos/toxicidade , Antibióticos Antineoplásicos/uso terapêutico , Dipeptídeos/toxicidade , Dipeptídeos/uso terapêutico , Neoplasias Hepáticas Experimentais/tratamento farmacológico , Nucleosídeos de Pirimidina/toxicidade , Nucleosídeos de Pirimidina/uso terapêutico , Sarcoma 180/tratamento farmacológico , Animais , Sobrevivência Celular/efeitos dos fármacos , DNA de Neoplasias/biossíntese , Dipeptídeos/isolamento & purificação , Fermentação , Humanos , Células KB , Neoplasias Hepáticas Experimentais/patologia , Camundongos , Proteínas de Neoplasias/biossíntese , Nucleosídeos de Pirimidina/isolamento & purificação , RNA Neoplásico/biossíntese , Streptomyces/crescimento & desenvolvimento , Ensaio Tumoral de Célula-TroncoRESUMO
The pyrimidine nucleoside, 1-beta-D-ribofuranosyl pyridine-2-one-5-carboxamide, is an anti inflammatory agent used in the treatment of adjuvant-induced arthritis. It is the 2-one isomer of 1-beta-D-ribofuranosyl pyridine-4-one 5-carboxamide, an unusual nucleoside isolated from the urine of patients with chronic myelogenic leukemia and an important cancer marker. Crystals of 1-beta-D-ribofuranosyl pyridine-2-one-5-carboxamide are monoclinic, space group C2, with the cell dimensions a = 31.7920(13), b = 4.6872 (3), c = 16.1838(11), beta = 93.071(3) degrees , V = 2408.2(2) A(3), D(calc) = 1.496 mg/m(3) and Z = 8 (two molecules in the asymmetric unit). The structure was obtained by the application of direct methods to diffractometric data and refined to a final R value of 0.050 for 1669 reflections with I >or= 3sigma. The nucleoside exhibits an anti conformation across the glycosidic bond (chi(CN) = -15.5 degrees , -18.9 degrees ), a C3 '-endo C2 '-exo [(3)(2)T] ribose pucker and g(+) across the C(4 ')-C(5 ') exocyclic bond. The amino group of the carboxamide group is distal from the 2-one and lacks the intramolecular hydrogen bonding found in the related 2-one molecule. Nuclear magnetic resonance studies shows also an anti conformation across the glycosidic bond but the solution conformation of the furanose ring is not the same as that found in the solid state.
Assuntos
Anti-Inflamatórios/síntese química , Artrite Experimental/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/urina , Modelos Moleculares , Nucleosídeos de Pirimidina/síntese química , Anti-Inflamatórios/química , Cristalografia por Raios X , Humanos , Ligação de Hidrogênio , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Nucleosídeos de Pirimidina/química , Nucleosídeos de Pirimidina/isolamento & purificaçãoRESUMO
A biologically guided fractionation from Lepista inversa (Scop.: Fr.) led to the isolation of clitocine, an exocyclic amino nucleoside. This compound and two mixtures of beta/alpha anomers (mixture A, 40:60 and mixture B, 80:20) were synthesized or isolated depending on the purification procedure. The beta anomer and clitocine mixtures A and B showed similar cytotoxic activities with IC50 values ranging from 20.5 to 42 nM in murine cancer cell lines (3LL and L1210) and from 185 to 578 nM in human cancer cell lines (DU145, K-562, MCF7, and U251). An in vivo study of mixture B was carried out on 3LL- and L1210-tumor-bearing mice. Clitocine solubilized in beta-hydroxypropylcyclodextrin and injected at concentrations of 0.5, 3, and 5 mg kg-1 did not significantly increase the survival rate and lifespan of 3LL-tumor-bearing mice. In contrast, clitocine showed antitumor activity on L1210-tumor-bearing mice with a significant increase in lifespan and a decrease in the development of ascites observed at 3 mg kg-1. The induction of apoptosis may be the basis of the antitumor activity of clitocine against L1210 as suggested by flow-cytometry analysis of cells treated in vitro.
Assuntos
Agaricales/química , Antineoplásicos/farmacologia , Nucleosídeos de Pirimidina/farmacologia , Animais , Antineoplásicos/isolamento & purificação , Divisão Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Cromatografia Líquida de Alta Pressão , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Espectroscopia de Ressonância Magnética , Camundongos , Nucleosídeos de Pirimidina/isolamento & purificaçãoRESUMO
The chromatographic behaviour of some natural and modified pyrimidine bases and nucleosides on an octadecyl stationary phase was studied. The retention and selectivity parameters of the separation of the compounds studied were derived on the basis of solvophobic theory. The mechanism of base and nucleoside interactions with the surface of the hydrocarbonaceous stationary phase is discussed. The best separation is observed at pH 3.5 for the bases and at pH 4.8-5.2 for the nucleosides. An increase in the solute surface tension results in an increased selectivity of separation. When the surface tension and the ionic strength of the mobile phase are not kept constant, there are considerable deviations in retention from that predicted by solvophobic theory.
Assuntos
Nucleosídeos de Pirimidina/isolamento & purificação , Pirimidinas/isolamento & purificação , Cromatografia Líquida de Alta Pressão/métodos , Concentração de Íons de Hidrogênio , Modelos Teóricos , Solventes , Relação Estrutura-AtividadeRESUMO
Chemical synthesis of DNA fragments with a defined sequence is extremely long and complex. A simpler alternative is isolation of fragments from partially hydrolysed DNA. Mixtures of sequence isomers of pyrimidine oligonucleotides were recently isolated on a preparative scale from hydrolysates of depurinated herring sperm DNA. These, however, could not be further separated by conventional techniques. Using high-performance liquid chromatography (HPLC), sequence isomers of pyrimidine di-, tri- and tetranucleotides were successfully separated and sequenced. In the present work, separation and sequencing of sequence isomers of pyrimidine pentanucleotides by HPLC was achieved, demonstrating the applicability of this particular approach in preparation of desired DNA fragments.